title: Sphingosine kinase-1 inhibition protects primary rat hepatocytes against bile salt-induced apoptosis.
abstract: Sphingosine kinases (SphKs) and their product sphingosine-1-phosphate (S1P) have been reported to regulate apoptosis and survival of liver cells. Cholestatic liver diseases are characterized by cytotoxic levels of bile salts inducing liver injury. It is unknown whether SphKs and/or S1P play a role in this pathogenic process. Here, we investigated the putative involvement of SphK1 and S1P in bile salt-induced cell death in hepatocytes. Primary rat hepatocytes were exposed to glycochenodeoxycholic acid (GCDCA) to induce apoptosis. GCDCA-exposed hepatocytes were co-treated with S1P, the SphK1 inhibitor Ski-II and/or specific antagonists of S1P receptors (S1PR1 and S1PR2). Apoptosis and necrosis were quantified. Ski-II significantly reduced GCDCA-induced apoptosis in hepatocytes (-70%, P<0.05) without inducing necrosis. GCDCA increased the S1P levels in hepatocytes (P<0.05). GCDCA induced [Ca(2+)] oscillations in hepatocytes and co-treatment with the [Ca(2+)] chelator BAPTA repressed GCDCA-induced apoptosis. Ski-II inhibited the GCDCA-induced intracellular [Ca(2+)] oscillations. Transcripts of all five S1P receptors were detected in hepatocytes, of which S1PR1 and S1PR2 appear most dominant. Inhibition of S1PR1, but not S1PR2, reduced GCDCA-induced apoptosis by 20%. Exogenous S1P also significantly reduced GCDCA-induced apoptosis (-50%, P<0.05), however, in contrast to the GCDCA-induced (intracellular) SphK1 pathway, this was dependent on S1PR2 and not S1PR1. Our results indicate that SphK1 plays a pivotal role in mediating bile salt-induced apoptosis in hepatocytes in part by interfering with intracellular [Ca(2+)] signaling and activation of S1PR1.
authors.affiliation: Dept. of Gastroenterology and Hepatology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands. Electronic address: g.karimian01@umcg.nl.
authors.forename: Golnar
authors.initials: G
authors.lastname: Karimian
authors.forename: Manon
authors.initials: M
authors.lastname: Buist-Homan
authors.forename: Martina
authors.initials: M
authors.lastname: Schmidt
authors.forename: Uwe J F
authors.initials: UJ
authors.lastname: Tietge
authors.forename: Jan Freark
authors.initials: JF
authors.lastname: de Boer
authors.forename: Karin
authors.initials: K
authors.lastname: Klappe
authors.forename: Jan Willem
authors.initials: JW
authors.lastname: Kok
authors.forename: Laurent
authors.initials: L
authors.lastname: Combettes
authors.forename: Thierry
authors.initials: T
authors.lastname: Tordjmann
authors.forename: Klaas Nico
authors.initials: KN
authors.lastname: Faber
authors.forename: Han
authors.initials: H
authors.lastname: Moshage
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001647:Bile Acids and Salts / Q000494:pharmacology*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002478:Cells, Cultured; D003902:Detergents / Q000494:pharmacology; D005765:Gastrointestinal Agents / Q000494:pharmacology; D005999:Glycochenodeoxycholic Acid / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D008246:Lysophospholipids / Q000378:metabolism; D008297:Male; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D060888:Real-Time Polymerase Chain Reaction; D049349:Receptors, Lysosphingolipid / Q000037:antagonists & inhibitors / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013110:Sphingosine / Q000031:analogs & derivatives / Q000378:metabolism; D013844:Thiazoles / Q000494:pharmacology; D000099157:Sphingosine Kinase
chemical_list: D001647:Bile Acids and Salts; D053148:Caspase 3; D003902:Detergents; D005765:Gastrointestinal Agents; D005999:Glycochenodeoxycholic Acid; D008246:Lysophospholipids; D017853:Phosphotransferases (Alcohol Group Acceptor); D012333:RNA, Messenger; D049349:Receptors, Lysosphingolipid; D013110:Sphingosine; D013844:Thiazoles; D000099157:Sphingosine Kinase; C525025:4-(4-(4-chloro-phenyl)thiazol-2-ylamino)phenol; C060506:sphingosine 1-phosphate
keywords: Calcium oscillation; Cholestatic liver disease; Glycochenodeoxycholic acid; Sphingosine kinase; Sphingosine-1 phosphate; Sphingosine-1 phosphate receptor